1 Weigh 500g of fresh ground beef without fat and fascia, add 1000mL of distilled water and 25mL of 1mol/L sodium hydroxide solution, stir and boil for 15min, fully cool, remove surface fat, clarify, filter, and add water to make up to 1000mL.Add various ingredients except meat scraps to correct the pH value.
2. The minced meat dregs are washed with water and dried to half dry. They are separately packed into 15mm × 150mm test tubes about 2-3 cm high. Each tube is filled withReduced iron powder0.1~0.2g or a little scrap iron.Replace the aboveLiquid mediumThe meat dregs shall be separately packed to about 1cm above the surface of the meat dregs in each tube.It is covered with melted vaseline or liquid paraffin for 0.3-0.4cm.Autoclave at 121 ℃ for 15min.
Detailed preparation
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Meat cooking medium (used for cultivating and preserving anaerobic bacteria)
(1) Fat free beef: take 500g of beef without fascia and fat, cut it into soybean sized particles, put it into a beaker containing 1000ml of distilled water, and boil it gently for about 1h.
(2) Filtration: filter with gauze, and then take a number of beef dregs and put them into a Hengate roll tube or ordinary test tube, with the loading capacity of about 15mm.Add beef extract peptone with PH7.4~7.6 in the test tubeLiquid medium10~12ml, and finally plugged with blackbutyl rubber Plug.
(3) Sterilization: insert a syringe needle on the isobutyl rubber plug, put it into the sterilization pot, and sterilize it at 121 ℃ for 20min.Remove the needle immediately after sterilization and plug the pipe plug.If the Hangate roller tubes of oxygen free culture medium are separately packed with oxygen free method, syringe needles can be dispensed with, but sterilization shall be carried out after the roller plugs are pressed tightly. The roller tubes thus prepared are called PRAS culture medium (or pre reducing anaerobic sterile culture medium), and oxygen removal is not necessary before use, but oxygen free is requiredAseptic manipulationTransfer or inoculation.
(4) Deaeration before use: ifAnaerobic mediumOxygen permeates during storage. Boil it for 10min in a pre positioned water bath to remove dissolved oxygen. After cooling under the saturation of high-purity nitrogen, sterilize it and avoid oxygen.
